Investigations will be directed at elucidating the role of guanosine triphosphate in microtubule structure and function. Tubulin will be purified from brain and neuroblastoma by affinity chromatography. Experiments designed to determine whether GTP is hydrolyzed during tubulin polymerization, depolymerization or other conformational alteration of microtubules will be performed. If GTP hydrolysis occurs, the non-hydrolyzable analogue, beta-gamma-methylene guanosine triphosphate should indicate the dependence of a structural change on the hydrolytic event. A knowledge of the localization and chemical nature of the GTP binding site(s) may indicate the manner in which the nucleotide influences tubulin structure. Thus, a series of group specific protein reagents will be allowed to react with tubulin. Peptides from tubulin preparations that appear to be modified at the GTP binding sites will be fingerprinted and identified. Previous observations suggest that other proteins may regulate and initiate tubulin polymerization; attempts will be made to show that such regulation actually occurs. In addition, the effect of phosphorylation on tubulin polymerization will be studied.